5-BROMO-2-(alpha-HYDROXYPENTYL)BENZOIC ACID SODIUM SALTS IN DIFFERENT CRYSTAL FORMS, AND PREPARATION METHOD THEREOF

ABSTRACT

The present invention discloses 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt in different crystal forms and the preparation methods thereof, and belongs to the field of pharmaceutical chemistry. Said different crystal forms of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt include: amorphous 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt, crystal form A of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt, and crystal form B of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt. The different crystal forms of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt obtained according to the present invention have better stability and water-solubility than the mixed forms of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt, thus is advantageous for pharmaceutical use. Moreover, the different crystal forms of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt possess much better therapeutic effect than 5-bromo-2-(α-hydroxypentyl)benzoic acid potassium salt.

REFERENCE TO RELATED APPLICATIONS

This application is a division of U.S. patent application Ser. No.15/660,395, filed Jul. 26, 2017, which is a division of U.S. patentapplication Ser. No. 14/905,199, filed Jan. 14, 2016, now U.S. Pat. No.9,902,682, issued Feb. 27, 2018, which is the U.S. national stageapplication of International PCT Application No. PCT/CN2014/081954,filed Jul. 10, 2014, which claims priority under 35 U.S.C. 119(b) toChinese Patent Application No. 201410313214.4, filed Jul. 3, 2014, andChinese Patent Application No. 20130299084.9, filed Jul. 17, 2013, theentire disclosures of which are incorporated herein by reference.

TECHNICAL FIELD

The present invention relates to 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt (BZP) in different crystal forms and preparation methodsthereof, and belongs to the field of pharmaceutical chemistry.

TECHNICAL BACKGROUND

Up to now, about half of drug molecules are present in the form of saltsand are administrated in the form of salts. By forming salts, physicaland chemical properties of drugs can be improved, such as solubility,dissolution rate, bioavailability, hygroscopicity, flowability, bulkdensity, stability, melting point, milling property, convenience forpreparation and purification, or the like. If the same drug hasdifferent crystal forms, the bioavailability of different crystal formswill be different. Moreover, the stability, flowability andcompressibility of different crystal forms will be different. Thus, suchdifferent physical and chemical properties will influence use of thedrug to some extent.

6-halo-3-butyl-1(3H)-isobenzofuranone compound has certain therapeuticeffect on ischemic cerebrovascular disease, and can promote recovery ofimpaired neural function in patients.6-halo-3-butyl-1(3H)-isobenzofuranone is substantially insoluble inwater, thus it is generally prepared as halo-2-(α-hydroxypentyl)benzoicacid, so as to improve the solubility. However,halo-2-(α-hydroxypentyl)benzoic acid is very unstable under roomtemperature and is easy to convert to6-halo-3-butyl-1(3H)-isobenzofuranone, therefore this is disadvantageousfor administration. It can be stored at low temperature (4° C.) forlimited storage period.

Chinese patent No. 01109795.7 discloses salts of2-(α-hydroxypentyl)benzoic acid and the preparation method and usethereof in the first time, which relates to salts of monovalent metalions, divalent metal ions and organic bases. It specifically disclosesthe salts of potassium, sodium, calcium, magnesium, zinc, aniline,benzylamine, morpholine, and diethylamine. Its specification alsodiscloses effect of the potassium salt on brain infarction area in ratswith local brain ischemia, effect of the potassium salt on plateletaggregation in rats, and protective effect of the potassium salt onischemia of isolated heart and arrhythmias following reperfusion inrats, thereby indicating the potassium salt produces beneficial effectin said experiments. Chinese patent No. 200410048268.9 and Chinesepatent No. 200610073077.7 respectively disclose the preparations andactivity tests of the corresponding chiral salts of2-(α-hydroxypentyl)benzoic acid. Chinese patent No. 200710054215.1discloses the synthesis and activity test of the halogenated compound6-halo-3-butyl-1(3H)-isobenzofuranone in the first time, wherein thehalogenated compound 6-halo-3-butyl-1(3H)-isobenzofuranone has obviouslybetter activity than butylphthalide. Chinese patent No. 200810230890. Xdiscloses the preparation method of halogenated2-(α-hydroxypentyl)benzoic acid salts compounds and pharmaceutical usethereof in the first time, which relates to salts of monovalent metalions, divalent metal ions and organic bases. It specifically disclosesthe salts of sodium, potassium, calcium, and benzylamine. Itsspecification also discloses the activity of the halogenated2-(α-hydroxypentyl)benzoic acid potassium salt for prevention andtreatment of heart and brain ischemic diseases, for improving heart andbrain circulatory disorders and antithrombotic effect, and the like,wherein activity of the halogenated 2-(α-hydroxypentyl)benzoic acidpotassium salt is better than butylphthalide,6-halo-3-butyl-1(3H)-isobenzofuranone, and 2-(α-hydroxypentyl)benzoicacid salt. However, the above patents do not relate to research ofcrystal forms of 5-halo-2-(α-hydroxypentyl)benzoic acid salts.Therefore, it is of practical importance to study different crystalforms of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt and toimprove pharmaceutical stability and water-solubility of said compound,thereby meeting the requirements for administration. In the meantime,the different crystal forms of 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt produce obviously better therapeutic effect than5-bromo-2-(α-hydroxypentyl)benzoic acid potassium salt (CNZL200810230890.X). Said better therapeutic effect has not been reportedby any literatures in the art until now.

DISCLOSURE OF THE INVENTION

In order to improve stability and water-solubility of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt, the presentinvention studied different crystal forms of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt, and providesdifferent crystal forms of 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt. Another object of the present invention is to provide themethods for preparing the crystal forms.

5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt according to thepresent invention has the following structural formula, which hasenantiomers:

Said different crystal forms of 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt include: amorphous 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt, crystal form A of 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt, and crystal form B of 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt.

The XRPD pattern of amorphous 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt generally has no obvious diffraction peak.

The present invention describes crystal form A of5-bromo-2-(α-hydroxypentyl) benzoic acid sodium salt, characterized inhaving XRPD pattern with the diffraction peaks at 2θ values of 5.60,16.46, 16.78, 18.77, 21.45, 32.28, 33.30, 33.49, 34.12, 36.14, 36.61,37.87, 40.24, 41.32, 43.76, 44.92, 45.18, 47.23, 55.12, 56.73, 57.12,62.78, wherein the error range of 2θ value is ±0.2.

The present invention describes crystal form B of5-bromo-2-(α-hydroxypentyl) benzoic acid sodium salt, characterized inhaving XRPD pattern with the diffraction peaks at 2θ values of 5.72,6.69, 9.93, 11.09, 11.84, 14.55, 16.42, 17.27, 17.80, 18.28, 19.86,20.98, 22.21, 23.43, 23.88, wherein the error range of 2θ value is ±0.2.

A method for preparing amorphous 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt: 6-bromo-3-butyl-1(3H)-isobenzofuranone and sodium hydroxideare weighed and placed in a crystallizer, to which tetrahydrofuran andwater are added, then the mixture is dissolved and reacted under a waterbath at 60° C.; after completion of the reaction, the reaction mixtureis distilled under reduced pressure to remove all the solvents to yieldamorphous 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt.

A method for preparing crystal form A of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt: amorphous5-bromo-2-(α-hydroxypentyl)benzoic acid is weighed, to which a solutionof NaOH in methanol is added, dissolved under ultrasound, and then thesolvent is volatilized slowly; after solids are precipitated, the solidsare dried under vacuum.

Method I for preparing crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt: amorphous5-bromo-2-(α-hydroxypentyl)benzoic acid is weighed, to whichtetrahydrofuran is added, and then the solution is stirred at roomtemperature; an aqueous solution of sodium hydroxide is slowly addeddropwise to the above-mentioned solution; after evaporation of solvents,solids are obtained, and dried under vacuum.

Method II for preparing crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt:6-bromo-3-butyl-1(3H)-isobenzofuranone and sodium hydroxide are weighedand placed in a flask; methanol is added to the flask, the resultingmixture is heated under reflux to react, and then methanol is evaporatedoff; ethyl acetate is added, the resulting mixture is shaken and theundissolved solids are filtrated off; ethyl acetate is evaporated off toyield solid compound, to which anhydrous diethyl ether is added todissolve, and the resulting solution is maintained standing overnightand is filtrated.

The inventiveness of the present invention is based on that5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt is prepared ascrystals. Since different crystal forms of the same compound aresignificantly different in appearance, solubility, melting point,dissolution rate, bioavailability and the like, pharmaceuticalstability, bioavailability and therapeutic effect will be affected. Thecrystal forms of 5-bromo-2-(α-hydroxypentyl)benzoic acid salt obtainedaccording to the present invention have better stability andwater-solubility than amorphous 5-halo-2-(α-hydroxypentyl)benzoic acid,thus are advantageous for pharmaceutical use.

DESCRIPTION OF FIGURES

FIG. 1 is DSC pattern of amorphous 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt;

FIG. 2 is XRPD pattern of amorphous 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt;

FIG. 3 is DSC/TGA pattern of crystal form A of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt;

FIG. 4 is XRPD pattern of crystal form A of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt;

FIG. 5 is DSC/TGA pattern of crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt;

FIG. 6 is XRPD pattern of crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt.

EXAMPLES

The following examples are used to help a skilled person in the artunderstand the present invention more deeply, but are not intended tolimit the present invention in any ways.

Example 1: Preparation of amorphous 5-bromo-2-(α-hydroxypentyl)benzoicAcid Sodium Salt

3.5 g of 6-bromo-3-butyl-1(3H)-isobenzofuranone and 520 mg of sodiumhydroxide were weighed and placed in a crystallizer, to which 40 mltetrahydrofuran and 8 ml water were added, then the mixture wasdissolved and reacted under a water bath at 60° C. After 3 hours, thereaction mixture was distilled under reduced pressure at 60° C. toremove all the solvents, to yield amorphous5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt.

Example 2: Preparation of Crystal Form A of5-bromo-2-(α-hydroxypentyl)benzoic Acid Sodium Salt

350 mg of amorphous 5-bromo-2-(α-hydroxypentyl)benzoic acid was weighed,to which 2 ml solution of NaOH in methanol was added, dissolved underultrasound, and then the solvent was volatilized slowly for 14 days.Solids were precipitated. The solids were dried under vacuum to yieldcrystal form A of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt.

Example 3: Preparation of Crystal Form B of5-bromo-2-(α-hydroxypentyl)benzoic Acid Sodium Salt (Method I)

1.15 g of amorphous 5-bromo-2-(α-hydroxypentyl)benzoic acid was weighedand placed in a 20-ml sample bottle with magnetically stirring at roomtemperature, to which 1 ml tetrahydrofuran was added. 120 mg of sodiumhydroxide was weighed and dissolved in 8 ml water, and then the NaOHsolution was slowly added into the above-mentioned solution dropwise.The resulting solution was subjected to rotary evaporation at 50° C.Solids were obtained, and dried under vacuum at room temperature toyield crystal form B of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodiumsalt.

Example 4: Preparation of Crystal Form B of5-bromo-2-(α-hydroxypentyl)benzoic Acid Sodium Salt (Method II)

20 g of 6-bromo-3-butyl-1(3H)-isobenzofuranone and 4.46 g of sodiumhydroxide were weighed and placed in a 500-ml round bottom flask. 300 mlmethanol was added to the flask, and the resulting mixture was heatedunder reflux to react. Then methanol was evaporated off 200 ml ethylacetate was added. After the resulting mixture was shook adequately, theundissolved solids were filtrated off. Ethyl acetate was evaporated offto yield viscous white solid compound, and 300 ml anhydrous diethylether was added to dissolve. The resulting solution was maintainedstanding overnight and a large amount of white solid precipitated. Themixture was filtrated to yield crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt.

Example 5: Effect of 5-bromo-2-(α-hydroxypentyl)benzoic Acid Sodium Salton Brain Infarction Volume and Brain Edema Volume in Rats with LocalBrain Ischemia

(1) Experimental Materials and Methods

Wistar rats (50% of rats is female and another 50% of rats is male);5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (BZP, in the form ofamorphous form, crystal form A and crystal form B, prepared according tothe present invention), prepared in double distilled water; TTC is inthe form of white powder, manufactured by Beijing Chemical Works.

Administration method: 5-bromo-2-(α-hydroxypentyl)benzoic acid sodiumsalt (BZP) was intravenously injected at one hour before MCAO, with thedosage of 12 mg/kg; 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt(BZP) was used to intragastric administration at one hour before MCAO,with the dosage of 20 mg/kg. 2-(α-hydroxypentyl)benzoic acid potassiumsalt (NBP-K) was used as positive control, and its dosage (calculated inmoles) is the same as that of 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt, that is, the dosage of intravenous injection is 9.6 mg/kg,and the dosage of intragastric administration is 15.9 mg/kg.

Preparation of MCAO model: Establish a focal brain ischemia-reperfusioninjury model by cerebral artery obstruction in rats with suture-occludedmethod. Rats were intraperitoneally injected with 10% chloral hydrate toanesthetize. Rats were fixed on back and the skin of neck center wasincised. The left common carotid artery, external carotid artery, andinternal carotid artery were subjected to blunt dissection. Thepterygopalatine artery was ligated, only remaining cranial trunk ofinternal carotid artery; a prepared nylon rope (burning diameter 0.3 mm)was inserted via external carotid artery incision into the bifurcationof the jugular vein, and was pushed along the cranial direction of theinternal carotid artery. Using the place of the bifurcation as startingpoint, after the rope was pushed forward about 17 mm, hand can feelforce of resistance, indicating that the head section of nylon rope hasbeen through the beginning of the middle cerebral artery. This indicatesone side of the middle cerebral artery occlusion model was completed.Sutured the incision, remained the end of nylon line out of the skin.For reperfusion, the nylon line was gently pulled out. If feel force ofresistance, this indicates the end of nylon line has returned to thetrunk of external carotid artery, thereby accomplishing reperfusion.

Determination of infarction volume: After being subjected to ischemiafor 2 hours and reperfusion for 24 hours, the heads of rats were cutoff. Forebrain was removed and cut into six pieces, each piece being 2mm. The pieces were stained with TTC, and incubated at 37° C. for 30minutes. Normal issues were stained to be red, but the infarcted issueswere stained to be white. Took photos with digital camera, andcalculated the percentage of the infarcted area relative to thespherical area of brain by Photoshop.

(2) Results

Experimental results of model group: brain infarction volume was53.42±4.65%, and brain edema volume was 13.66±3.46%.

The effect of intravenous injection and intragastric administration of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (BZP) on braininfarction volume in rat MCAO model and the effect of intravenousinjection and intragastric administration of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (BZP) on brain edemavolume in rat MCAO model were as follows:

Effect of intravenous injection of 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt (BZP) on brain infarction volume in rat MCAO model

Inhibition of Infarction infarction Dosage volume volume Drug (mg/kg) n(%) (%) Pseudo-surgery 0 0 group Model group 10 53.42 ± 4.65    BZP(amorphous 12 10 3.46 ± 2.94 aa 93.52 form) BZP (crystal form 12 10 4.54± 1.73 aa 91.50 A) BZP (crystal form 12 10 4.76 ± 1.45 aa 91.09 B) NBP-K9.6 10 24.35 ± 4.32 aa  54.42 aa: P < 0.01, as compared with the modelgroup; ** P < 0.01, as compared with the NBP-K group.

Effect of intravenous injection of 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt (BZP) on brain edema in rat MCAO model

Inhibition Dosage Edema of edema Drug (mg/kg) n (%) (%) Pseudo-surgery 00 group Model group 10 13.66 ± 3.46 BZP (amorphous 12 10    2.56 ± 1.63aa 88.58 form) BZP (crystal form 12 10     3.24 ± 4.11 aa * 81.28 A) BZP(crystal form 12 10  4.20 ± 2.03a 78.71 B) NBP-K 12 10 12.38 ± 1.29 9.37aa: P < 0.01, as compared with the model group; * P < 0.05, as comparedwith the NBP-K group; ** P < 0.01, as compared with the NBP-K group.

Effect of intragastric administration of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (BZP) on braininfarction volume in rat MCAO model

Inhibition of Infarction infarction Dosage volume volume Drug (mg/kg) n(%) (%) Pseudo-surgery 0 0 group Model group 10 53.42 ± 4.65    0 BZP(amorphous 20 10 5.59 ± 2.35 aa 89.05 form) BZP (crystal form 20 10 7.13± 2.98 aa 86.65 A) BZP (crystal form 20 10  7.13 ± 2.98 aa** 86.03 B)NBP-K 15.9 10 22.28 ± 2.65 aa  58.3 aa: P < 0.01, as compared with themodel group; **P < 0.01, as compared with the NBP-K group.

Effect of intragastric administration of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (BZP) on brain edemain rat MCAO model

Inhibition Dosage Edema of edema Drug (mg/kg) n (%) (%) Pseudo-surgery 00 group Model group 10 13.66 ± 3.46    BZP (amorphous 20 10 2.24 ±1.66^(aa)** 88.36 form) BZP (crystal form 20 10  2.7 ± 1.30^(aa)** 87.55A) BZP (crystal form 20 10 2.87 ± 1.56^(aa)** 85.09 B) NBP-K 15.9 1011.04 ± 1.48^(aa)**  42.65 ^(aa)P < 0.01, as compared with the modelgroup; * P < 0.05, as compared with the NBP-K group; **P < 0.01, ascompared with the NBP-K group.

(3) Conclusion

5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt can obviously reducebrain tissue injury induced by obstruction of cerebral arteries, reducebrain infarction volume and reduce brain edema volume in rats. At thesame dosage, the activity of 5-bromo-2-(α-hydroxypentyl)benzoic acidsodium salt is obviously better than 2-(α-hydroxypentyl)benzoic acidpotassium salt, and the activity of 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt is also better than butylphthalide and halogenated2-benzo[c]furanone known in the art.

Example 6: Results of Primary Toxicity Study of5-bromo-2-(α-hydroxypentyl)benzoic Acid Sodium Salt

Data of acute toxicity and sub-acute toxicity of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt BZP (in amorphousform)

BZP NBP-K Items ig iv ig iv LD50 Mouse 927 323 880 303 (mg/Kg) Rat 726246 623 230 Sub-acute Rat >20 >12 >20 >12 (mg/Kg) Note: ig 20 mg/kg; iv12 mg/kg.

Example 7: Sub-Acute Toxic Effect of 5-bromo-2-(α-hydroxypentyl)benzoicAcid Sodium Salt (in Amorphous Form) on Liver and Kidney Functions inRats

Groups Control ig iv Items Sex group (20 mg/kg) (12 mg/kg) ALT F 43.38 ±6.69 35.10 ± 8.61  38.00 ± 9.82  AST F 219.40 ± 55.38 168.00 ± 17.19*202.40 ± 68.12  M  5.30 ± 0.87 4.92 ± 0.75 5.34 ± 1.33 AST/ALT F  246.20± 129.61 135.29 ± 48.13* 184.80 ± 68.59  M  4.35 ± 0.05  2.74 ± 1.62*3.96 ± 1.20 Total Proteins F 48.52 ± 2.46 46.63 ± 1.07  49.00 ± 0.79 (TP) M 41.70 ± 2.04 40.26 ± 15.78 53.40 ± 8.69* Albumin F 33.52 ± 1.9832.43 ± 1.14  33.96 ± 1.71  (ALB) M 27.74 ± 1.38 28.98 ± 7.34  36.90 ±6.72* Total Bilirubin F  2.58 ± 0.63 2.10 ± 0.40 2.72 ± 0.67 (TBIL) M 5.20 ± 3.88  2.24 ± 1.50**  3.10 ± 1.23* Blood urea nitrogen F  3.69 ±0.46 3.80 ± 0.59 4.00 ± 0.43 (BUN) M  4.54 ± 0.72  2.76 ± 1.09**  3.46 ±0.72* Creatinine F 59.80 ± 5.02 57.50 ± 3.11  55.40 ± 2.30  (CRE) M60.00 ± 7.25  38.80 ± 25.62* 66.60 ± 15.09 Creatinine kinase F  924.20 ±445.66 776.00 ± 437.68 636.80 ± 536.05 (CK) M 634.50 ± 37.48  938.60 ±553.43* 762.00 ± 351.72 Note: *P < 0.05, **P < 0.01, as compared withthe same gender of control group.

Example 8: Sub-Acute Toxic Effect of 5-bromo-2-(α-hydroxypentyl)benzoicAcid Sodium Salt (in Amorphous Form) on Blood Routine Tests

Groups Control ig iv Items Sex group (20 mg/kg) (12 mg/kg) RBC F 6.97 ±1.80 4.51 ± 3.30 6.95 ± 1.06 M 7.67 ± 0.53 7.61 ± 0.28 7.86 ± 0.40 HGB F126.60 ± 30.22  117.00 ± 19.70  125.60 ± 16.68  M 140.33 ± 5.69  134.00± 19.13  141.40 ± 7.20  PLT F 1129.75 ± 109.48  1146.00 ± 74.22  1075.00± 41.70  M 1118.67 ± 53.26  1258.20 ± 281.33  1087.20 ± 56.34  WBC F5.50 ± 2.49 4.44 ± 2.24 5.20 ± 1.53 M 4.67 ± 2.22 5.18 ± 0.71 5.66 ±2.23 Neutrophil F 22.20 ± 7.96  28.44 ± 10.20 21.44 ± 2.74  (NEU) M17.23 ± 5.66  23.88 ± 5.48  22.16 ± 4.14  Lymphocyte F 71.26 ± 7.86 66.45 ± 11.13 73.62 ± 2.16  (LYM) M 73.53 ± 9.11  70.84 ± 3.99  72.50 ±4.66  Monocyte F 4.56 ± 2.12  2.05 ± 1.74** 3.62 ± 0.63 (MON) M 7.20 ±323   4.93 ± 1.91*  3.80 ± 0.70* Eosinophil F 0.70 ± 0.50  0.23 ± 0.12**0.76 ± 0.50 (EOS) M 0.63 ± 0.45 0.56 ± 0.50 0.60 ± 0.29 Basophil F 1.28± 0.85 1.43 ± 1.17  0.56 ± 0.15** (BAS) M 1.40 ± 0.20  0.62 ± 0.25* 0.40 ± 0.12** Note: *P < 0.05, **P < 0.01, as compared with the samegender of control group.

Example 9: Sub-Acute Toxic Effect of 5-bromo-2-(α-hydroxypentyl)benzoicAcid Sodium Salt (in Amorphous Form) on Blood Coagulation Functions inRats

Groups Control ig iv Items Sex group (20 mg/kg) (12 mg/kg) APTT F 25.6 ±2.3 22.8 ± 5.9 22.3 ± 8.5 (sec) M 25.5 ± 3.2 26.5 ± 3.2 23.7 ± 2.1 PT F18.0 ± 2.1 17.5 ± 2.2 18.8 ± 3.3 (sec) M 18.4 ± 1.8 16.4 ± 3.4 17.5 ±2.0 FIB F  7.31 ± 0.51  7.22 ± 0.97  6.63 ± 0.95 (g/l) M  6.66 ± 1.08 7.03 ± 0.29  6.28 ± 0.55 TT F 26.3 ± 2.5 27.3 ± 2.4 27.5 ± 2.8 (sec) M28.5 ± 1.7 26.9 ± 5.9 27.1 ± 3.8

Example 10: Effect of 5-bromo-2-(α-hydroxypentyl)benzoic Acid SodiumSalt (in Amorphous Form) on Body Weight of Rats

Day 1 Day 6 Day 11 Day 16 Day 21 Day 24 Groups Sex BW BW ΔW (%) BW ΔW(%) BW ΔW (%) BW ΔW (%) BW ΔW (%) Control M 178 ± 6  207 ± 8 16 ± 5  253± 4  22 ± 3 295 ± 10 66 ± 6  330 ± 20 85 ± 9  341 ± 24 92 ± 10  F 172 ±6  18

 ± 6 9 ± 2 216 ± 10 15 ± 3 260 ± 6  37 ± 5  258 ± 11 50 ± 6  259 ± 10 52± 7  ig group M 183 ± 8  210 ± 8 15 ± 3  245 ± 12 17 ± 5 280 ± 18 53 ±14 320 ± 18 75 ± 14  337 ± 18 84 ± 14  F 179 ± 12  192 ± 16 7 ± 4 205 ±14  7 ± 2* 224 ± 11 25 ± 4* 230 ± 15 29 ± 6** 235 ± 12 31 ± 5** iv groupM 179 ± 13 204 ± 7 14 ± 6  238 ± 6   14 ± 2* 270 ± 6  51 ± 10 296 ± 8 66 ± 15* 304 ± 10 71 ± 16* F 182 ± 13  195 ± 12 7 ± 2 200 ± 15   3 ± 2**212 ± 14  21 ± 7** 223 ± 20 23 ± 6** 221 ± 19 21 ± 6** Note: ΔW (%)means the percentage of increase of body weight of rats at Day X ascompared with the body weight at Day 1. *P < 0.05, as compared with thepercentage of increase of body weight of the same gender in controlgroup, **P<0.01, as compared with the percentage of increase of bodyweight of the same gender in control group. Note: ig: 20 mg/kg; iv: 12mg/kg.

indicates data missing or illegible when filed

Example 11

Comparison of stability and solubility of amorphous5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (BZP, in the form ofamorphous form), crystal form A of 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt (BZP crystal form A) and crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (BZP crystal form B)with 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt (a mixture, CNZL200810230890.X) in water and methanol solution

Solubility Solubility in water Stability in methanol Stability Drag(g/mL) (hours) (g/mL) (hours) BZP (amorphous) 2.98 >24 1.89 >24 BZP(Form A) 2.36 >20 1.80 >24 BZP (Form B) 2.43 >24 1.63 >24 BZP (mixture)2.13 >12 1.58 >12

What is claimed is:
 1. Crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt, defined by a XRPDpattern with the diffraction peaks at 2θ values of 5.72, 6.69, 9.93,11.09, 11.84, 14.55, 16.42, 17.27, 17.80, 18.28, 19.86, 20.98, 22.21,23.43, and 23.88, wherein the error range of 2θ value is ±0.2.
 2. Amethod for preparing the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1, themethod comprising: dissolving amorphous5-bromo-2-(α-hydroxypentyl)benzoic acid in a tetrahydrofuran solvent,stirring the resulting solution at room temperature, and adding anaqueous solution of sodium hydroxide dropwise to the solution; whereinafter evaporation of the tetrahydrofuran and water solvents, theresulting solids are dried under a vacuum.
 3. A method for preparing thecrystal form B of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt ofclaim 1, the method comprising: weighing and placing6-bromo-3-butyl-1(3H)-isobenzofuranone and sodium hydroxide in a flask,adding methanol to the flask, heating the resulting mixture under refluxto react the contents, evaporating and removing the methanol, addingethyl acetate, shaking the resulting mixture and filtering and removingthe undissolved solids, evaporating and removing the ethyl acetate toyield a solid compound, adding anhydrous diethyl ether to dissolve thesolid compound, and filtering the resulting solution after standingovernight.
 4. A method of reducing brain tissue injury induced byobstruction of cerebral arteries comprising administering the crystalform B of 5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt ofclaim
 1. 5. A method of reducing brain infarction volume comprisingadministering the crystal form B of 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt of claim
 1. 6. A method of reducing brain edema volumecomprising administering the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim
 1. 7. Amethod of preventing and treating heart and brain ischemic diseasescomprising administering the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim
 1. 8. Amethod of improving heart and brain circulatory disorders comprisingadministering the crystal form B of 5-bromo-2-(α-hydroxypentyl)benzoicacid sodium salt of claim
 1. 9. A method of producing antithromboticeffects comprising administering the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim
 1. 10. Themethod of claim 4 wherein administration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintravenous administration.
 11. The method of claim 4 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintragastric administration.
 12. The method of claim 5 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintravenous administration.
 13. The method of claim 5 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintragastric administration.
 14. The method of claim 6 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintravenous administration.
 15. The method of claim 6 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintragastric administration.
 16. The method of claim 7 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintravenous administration.
 17. The method of claim 7 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintragastric administration.
 18. The method of claim 8 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintravenous administration.
 19. The method of claim 8 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintragastric administration.
 20. The method of claim 9 whereinadministration of the crystal form B of5-bromo-2-(α-hydroxypentyl)benzoic acid sodium salt of claim 1 comprisesintravenous administration or intragastric administration.